Although Escherichia coli is considered part of the normal bowel flora of healthy individuals, the microorganism is also recognized as an important pathogen in human diseases and conditions, including urinary tract infections, meningitis, and diarrhea. Therefore, a highly sensitive and specific method that can rapidly identify E. coli in biological samples would be useful.
The invention is based on the discovery that a highly sensitive and specific E. coli agglutination assay can be performed using antibodies against total urea-extracted E. coli antigens. Surprisingly, the sensitivity of the assay is dramatically reduced when using antibodies against specific E. coli antigens, even though such a strategy proved useful in agglutination assays for other bacterium (see, e.g., U.S. Pat. No. 5,418,140). Consequently, the invention relates in part to the unexpectedly superior performance of the assay using antibodies against total urea-extracted antigens, rather than specific surface antigens.
Accordingly, the invention features a method for detecting Escherichia coli in a sample by providing a sample suspected of containing Escherichia coli; contacting the sample with beads coated with a mixture of antibodies or anti-serum specific to one or more surface antigens of Escherichia coli, each of the one or more surface antigens having a molecular weight of 21xc2x12 KDa, 26xc2x12 KDa, 31xc2x12 KDa, 36xc2x12 KDa, 38xc2x12 KDa, 67xc2x12 KDa, or 77.8xc2x12 KDa; and observing agglutination of the beads, where the presence of the agglutination indicates the presence of Escherichia coli in the sample. The method can further include determining whether a microorganism in the sample produces indole or ferments lactose, both of which are characteristic of E. coli. 
The invention further features a kit for testing whether E. coli is present in a sample. The kit contains beads and a mixture of antibodies or antisera specific to one or more surface antigens of Escherichic coli, wherein each of the one or more surface antigens has a molecular weight selected from the group consisting of 21xc2x12 KDa, 26xc2x12 KDa, 31xc2x12 KDa, 36xc2x12 KDa, 38xc2x12 KDa, 67xc2x12 KDa, and 77.8xc2x12 KDa. The kit can also include (1) a composition containing eosin-methylene blue for determining whether a microorganism ferments lactose, (2) a composition containing p-dimethylaminocinnamaldehyde for determining whether a microorganism produces indole, (3) a positive control reagent that is expected to agglutinate the beads coated with the mixture of antibodies, or (4) a negative control reagent that is not expected to agglutinate the beads coated with the mixture of antibodies.
As used herein, xe2x80x9ctotal urea-extracted Escherichia coli antigensxe2x80x9d means the antigens collected from the supernatant of a centrifuged mixture containing E. coli bacteria incubated in an aqueous mixture containing about 4 M urea.
Other features or advantages of the present invention will be apparent from the following detailed description, and also from the claims.
The invention relates to methods of detecting E. coli in a sample, such as a biological sample (e.g., a serum or cerebral spinal fluid sample) using an agglutination assay. While agglutination assays have been used to detect various species of bacteria in samples, the moieties attached to the beads and useful for agglutination are different, depending on the species to be detected (see, e.g., Chang et al., J. Food Protect. 57:31-36, 1994; and Chang et al., J. Food Protect. 56:759-762, 1993). Indeed, it is thus far uncertain which moieties are needed to detect a different species of bacterium.
The present invention is based in part on the identification of the particular moiety required for a highly sensitive and specific agglutination assay for detection of E. coli. As shown in the example below, the E. coli agglutination assay is not specific enough for clinical applications unless the beads are coated (e.g., by standard covalent or non-covalent methods) with a mixture of antibodies specific for total urea-extracted E. coli antigens. Such a mixture of antibodies can be produced by immunizing a vertebrate with total urea-extracted E. coli antigens and harvesting the antibodies produced by the animal, using standard techniques such as the ones described in the example below.
Without further elaboration, it is believed that one skilled in the art can, based on the above disclosure and the agglutination assays described below, utilize the present invention to its fullest extent. The following example is to be construed as merely illustrative of how one skilled in the art can isolate total urea-extracted E. coli antigens, immunize an animal with such antigens, and harvest antibodies produced by the animal, and are not limitative of the remainder of the disclosure in any way. Any publications cited in this disclosure are hereby incorporated by reference.